Antimicrobial and sporicidal composition

ABSTRACT

Germicidal compositions with enhanced activity towards killing microbiological spores and vetgetative cells comprising certain quaternary ammonium compounds (QACs), phenolic compounds, monohydric alcohols, hydrogen peroxide, iodine, triclocarban, triclosan or combinations thereof with one or more spore coat opening agents. The invention also provides for the application of the germicidal compositions to animate and inanimate surfaces to help kill germs and protect against the risk of infection from bacteria, molds, yeasts, fungi, viruses, and microbiological spores.

PRIORITY DATA

This application is a continuation of U.S. patent application Ser. No.10/346,216, filed on Jan. 16, 2003, which claims the benefit of U.S.Provisional Application No. 60/350,416 filed on Jan. 18, 2002, each ofwhich is incorporated herein by reference.

BACKGROUND OF THE INVENTION

1. Field of the Invention

The present invention relates generally to germicidal disinfectingcompositions. More particularly, the present invention relates to anovel concept for germicidal disinfecting compositions that areeffective in killing both vegetative microorganisms and microbiologicalspores in a single composition.

2. Related Art

Microbiological spores are among the most resilient forms of dormantlife known to man. Although formed by many different types ofmicrobiological organisms, the most commonly studied spores are fromvarious Bacillus or Clostridium species. In particular, Bacillus sporesare amazingly resistant to common sterilizing techniques. For example,most vegetative bacteria die quickly when subjected to temperatures inexcess of 80° C., but bacterial spores often survive boiling water at100° C. for two hours or more. Spores have survived for 20 years in 70%alcohol solutions and can survive one million rems of radiation. Dryinghas little effect on spores, as demonstrated by spores surviving in theintestines of Egyptian mummies for thousands of years. [I. EdwardAlcamo, (2001) Fundamentals of Microbiology, 6^(th) ed., Jones andBartlett Publishers, Sudbury, Mass.].

Spores are formed by bacteria when nutritional resources are low. Theresulting spores have certain compartments and layers. The central core,referred to as the protoplast, contains soluble enzymes and geneticmaterial. The spore protoplast is surrounded by a spore membrane thatfunctions as a boundary for the spore protoplast, as a regulator of cellpermeability, and as a structural matrix for certain enzymes. A germcell wall is immediately external to the spore membrane, composedprimarily of peptidoglycan. The next layer is the spore cortex, alsocomposed of modified peptidoglycan. Many of the properties of theprotective cortex layer are attributed to this component such as theregulation of spore size and permeability. The spore coat represents30-60% of the dry spore weight, contains approximately 80% of the totalspore proteins, and occupies about 50% of the spore volume. The sporecoat is extremely resistant to a variety of drastic chemical treatments,suggesting that the coat plays an important role in spore resistance.The exposporium, or outermost spore layer varies in structure andcomposition from species to species of bacteria.

Spores remain dormant until germination is triggered by externalstimuli. Many types of spores are activated by heat or by certainchemical substances such as calcium dipicolinate and amino acids, mostparticularly L-alanine. Activation is the initial event in germination,an irreversible process that converts dormant spores into metabolicallyactive ones within a short period of time. Initiation of germination isfollowed rapidly by a number of degradative changes in the spore. Theprotective coating around the spore begins to open and becomes morepermeable to water, causing the germinated spores to swell. Reducingagents that reduce disulfide cystine linkages in proteins and enzymesthat attack peptidoglycan linkages also weaken the spore coat andincrease permeability of the spore. As germination proceeds, the sporesshed their outer layers, allowing the young, vegetative cells to emerge,elongate, and divide. Development of a vegetative cell from a germinatedspore is called outgrowth.

A safe and effective topical skin care formulation capable of killingmicrobiological spores and preventing their subsequent development intoinfectious, vegetative cells is desired and needed. Furthermore, it isalso desired that the formulation be gentle to skin, provide acosmetically pleasing feel and assist in gently moisturizing the skin,such that the user will gain benefit from both the microbiocidalactivity and improved skin health through repetitive application.

SUMMARY OF THE INVENTION

It has been recognized that it would be advantageous to developgermicidal composition with enhanced microbiocidal activities forkilling microbiological spores, microbiological outgrowth from spores,and vegetative microorganisms. The invention provides a germicidalcomposition with enhanced microbiocidal activities for killingmicrobiological spores, microbiological outgrowth from spores, andvegetative microorganisms and a method of use thereof.

In a first embodiment, the composition of the present inventioncomprises at least one quaternary ammonium compound selected from thegroup consisting of a quaternary ammonium compound containing a C₁₈₋₃₀alkyl substituent, benzethonium chloride, benzethonium bromide,benzethonium iodide, methylbenzethonium chloride, methylbenzethoniumbromide, and methylbenzethonium iodide. Preferably, the quaternaryammonium compound is present in a concentration of about 0.01% to 10%,and a more preferably in a concentration of about 0.05% to 8%, or anoptimal concentration of about 0.08% to 5%.

In a second embodiment, the composition comprises an antibacterial agentand a spore coat opener.

In this embodiment, the antibacterial agent is a member selected fromthe group consisting of a quaternary ammonium compound, a phenoliccompound, a monohydric alcohol, hydrogen peroxide, iodine, triclocarban,triclosan or combinations of such antibacterial agents.

The concentration of the antibacterial agent may vary depending upon theclass selected. The quaternary compound may be present in aconcentration of about 0.01% to 10%, or more preferably in aconcentration of about 0.05% to 8%, or in an optimal concentration ofabout 0.08% to 5%. The phenolic compound is selected from the groupconsisting of phenol, para-chlorometaxylenol, cresol, resorcinol, andhexylresorcinol and may be present in a concentration of about 0.01% to10%, or more preferably in a concentration of about 0.01% to 5%, or inan optimal concentration of about 0.05% to 1%. The monohydric alcohol isselected from the group consisting of ethyl alcohol and isopropylalcohol, and may be present in a concentration of about 20% to 95%, ormore preferably in a concentration of about 20% to 70%, or in an optimalconcentration of about 25% to 70%. The hydrogen peroxide may be presentin a concentration of about 1% to 10%, or more preferably in aconcentration of about 1% to 5%, or in an optimal concentration of about2% to 4%. The iodine may be present in a concentration of about 0.1% to10%, or more preferably in a concentration of about 0.5% to 8%, or in anoptimal concentration of about 1% to 5%. The triclocarban or triclosanmay be present in a concentration of about 0.01% to 5.0%, or morepreferably in a concentration of about 0.05% to 3%, or in an optimalconcentration of about 0.05% to 2%.

As noted, in the second embodiment, the composition of the presentinvention also contains at least one spore coat opener selected from thegroup consisting of an amino acid, a metal chelation agent, a reducingagent, an emulsifier/surface-active agent or a mixture of any or all ofthese agents.

Preferably, the amino acid is selected from the group consisting ofalanine, glycine, valine, leucine, isoleucine, proline, serine,threonine, methionine, cysteine, taurine, tyrosine, phenylalaine,tryptophan, asparagine, glutamine, aspartic acid, glutamic acid, lysine,arginine, and histidine, and combinations thereof including their freeacid forms and their salts. The amino acid is present in a concentrationof about 0.0001% to 5%, or more preferably in a concentration of about0.001% to 2%, or in an optimal concentration of about 0.005% to 1%.

The metal chelation agent is selected from the group consisting of EDTA,EGTA, picolinic acid, nicotinic acid, niacin, citric acid, ascorbicacid, glutamic acid, and fulvic acid, and combinations thereof includingtheir free acid forms and their salts. The chelation agent is present ina concentration of about 0.001% to 5%, or more preferably in aconcentration of about 0.01% to 2%, or in an optimal concentration ofabout 0.05% to 1%.

The reducing agent is selected from the group of substances orcombinations of substances that are capable of reducing disulfide bonds,including ascorbic acid, quinone, polyphenol with up to hundreds ofpolymeric subunits including but are not limited to: phenol-richpolymers of flavonoids, gallic acid, ellagic acids and their respectivecarbohydrate esters, salts and derivatives, and proanthocyanidin,including their free acid forms. The reducing agent is present in aconcentration of about 0.001% to 2%, or more preferably in aconcentration of about 0.001% to 1%, or in an optimal concentration ofabout 0.005% to 0.5%.

The emulsifier/surface-active agent is selected from the groupconsisting of C₁₂₋₂₄ monohydric alcohols, C₂₋₁₈ monohydric alcoholethoxylates, C₈₋₂₄ alkylamine oxides, silicone copolyols, the quaternaryammonium compounds behentrimethylammonium, babassuamindopropalkonium,benzethonium, benzalkonium, a C₈₋₂₄ mono-alkylquaternary ammonium, aC₈₋₂₄ di-alkylquaternary ammonium and the like, combined with theirrespective anionic salt counterparts chosen from the group of halogenssuch as chloride, bromide, or iodide, methosulfate, ethosulfate,acetate, lactate, propionate, and the like. Theemulsifier/surface-active agent is present in a concentration of about0.1% to 25%, or more preferably in a concentration of about 0.5% to 15%,or in an optimal concentration of about 1% to 10%. Combinations of twoor more emulsifier/surface-active agents may also be utilized.

Additional features and advantages of the present invention will beapparent from the detailed description which follows, taken inconjunction with the accompanying drawings, which together illustrate,by way of example, features of the invention.

BRIEF DESCRIPTION OF THE DRAWINGS

FIG. 1 illustrates generalized chemical structures of quaternaryammonium chloride compounds with antimicrobial activity.

FIG. 2 is graphic illustration of the test results of spore-inactivatingcompositions of the present invention.

DETAILED DESCRIPTION

Before the present composition and methods of making and using thereofare disclosed and described, it is to be understood that this inventionis not limited to the particular configurations, as process steps andmaterials may vary somewhat. It is also intended to be understood thatthe terminology employed herein is used for the purpose of describingparticular embodiments only and is not intended to be limiting since thescope of the present invention will be limited only by the appendedclaims and equivalents thereof. It must be noted that, as used in thisspecification and the appended claims, the singular forms “a,” “an,” and“the” include plural referents unless the context clearly dictatesotherwise.

“germ” refers to a small mass of living substance capable of developinginto an organism or one of its parts; a microorganism causing disease.

“germicide” refers to an agent that destroys or deactivates germs.

Reference will now be made to the exemplary embodiments illustrated andspecific language will be used herein to describe the same. It willnevertheless be understood that no limitation of the scope of theinvention is thereby intended. Alterations and further modifications ofthe inventive features illustrated herein, and additional applicationsof the principles of the invention as illustrated herein, which wouldoccur to one skilled in the relevant art and having possession of thisdisclosure, are to be considered within the scope of the invention.

This invention relates to germicidal compositions with enhancedgermicidal activities for killing microbiological spores, vegetativecells that arise from spores, bacteria, viruses, molds, yeasts, fungi,and the like. Expanded activity of germicidal compositions is providedby the inclusion of agents such as amino acids, metal chelation agents,reducing agents, or surface-active agents that open or weaken sporecoats thereby increasing permeability to the germicidal agents.Therefore, the novel combination of spore coat opening agents withantimicrobial agents expands their traditional activities to includesignificantly enhanced and expanded activity against spores.

The invention also provides for the application of the germicidalcompositions to animate and inanimate surfaces to help prevent the riskof infection from bacteria, molds, yeasts, viruses, fungi, andmicrobiological spores.

The invention describes germicidal compositions that contain one or moreantimicrobial and sporicidal active ingredients. In particular,quaternary ammonium compounds (QACs) are well known for theirantibacterial action against certain types of vegetative cells. Animportant aspect of this invention is the application of specificclasses of QACs as sporicidal agents per se. Certain surface-active QACsexhibit sporicidal activity, such as linear alkyl and benzylalkoniumchlorides, dialkyldimethylammonium chlorides, and the rather unique QAC:benzethonium chloride (BEC). The chemical structure of BEC represents acreative departure from the relatively simple alkyl groups substitutedon the quaternary nitrogen. The distinguishing feature of this rathercomplex molecule is best described by BEC's IUPAC name,N,N-Dimethyl-N-[2-[2-[4-(1,1,3,3-tetramethylbutyl)phenoxy]ethoxy]ethyl]benzene-methanaminiumchloride. As seen in FIG. 1, this complex nitrogen substituent containsbranched hydrocarbons, phenoxy, and ethoxy components. Ethoxylationincreases detergency and the complex nature of this group enhances thespectrum of activity when compared to more traditional QACs. The uniquechemical character of BEC establishes it as a very different class ofQACs. BEC's sporicidal activity, or its method of use in destroyingmicrobiological spores has not previously been reported.

An important aspect of the present invention is to combine ingredientsinto a formulation that will help to open up the protective sporecoatings, so as to facilitate the entry of antimicrobial ingredientsinto the inner core of the spores where they can interrupt normal sporefunctions, thus killing or inactivating the spore more effectively thanwithout such additives. Furthermore, the new formulation may be in theform of an oil-in-water emulsion that is very stable in both freezingand elevated storage temperatures which enhances product stability.

Another aspect of the invention is to insure that the formulation offersa variety of ingredient combinations to assist those skilled in the artin developing combinations of these ingredients into desirablegermicidal products.

As noted above, certain QACs are both antimicrobial and sporicidal.Other QACs are antimicrobial only.

Low molecular weight alcohols are excellent antimicrobial activeingredients. However, due to the very different anatomy and function ofspores as compared to vegetative forms of infectious organisms, alcoholsare not sporicidal. In fact, aqueous-alcoholic solutions are the storagemedium of choice for bacterial spores. Spores survive intact for yearswhen stored in aqueous-ethanol solutions.

Conversely, aldehydes, phenols, parabens, elemental halogens, and ozonecan inhibit various steps of the spore germination process followingactivation during subsequent outgrowth. To this point in time, only lowmolecular weight aldehydes have been shown to demonstrate sporicidalactivity against dormant spores, prior to their activation. It is knownthat combining one or more of these sporicidal agents often results in amore active composition. For example combinations of QACs withdialdehydes has been a particularly active combination in coldsterilizers for inanimate surfaces.

One particularly important aspect of the present invention relates tothe combination of active sporicides with other ingredients that are notindependently sporicidal resulting in a composition that workssynergistically with enhanced sporicidal activity. Combining sporicidalagents with substances that exhibit spore-activating properties for thepurpose of enhancing sporicidal activity provides a novel aspect of theinvention. Three such useful groups of spore-activating ingredients areamino acids, divalent metal chelates, and mild reducing agents. Unlikethe sporicidal agents, these latter ingredients stimulate the dormantspore, through various mechanisms, to initiate a breakdown of theprotective coating around the spore, causing a weakening of the sporecoating. The resulting gaps and channels and increased permeability inthe spore coating allow penetration of biocidal agents into the sporecore and inner membranes, where they can act to destroy the spore moreeffectively.

The invention also makes use of a similar effect on spore coat weakeningthat can be achieved by incorporating an emulsion of oil and water toweaken and open the spore coat, making the spore more susceptible todamage. The premature opening of the spore coat caused by contact withsuch an emulsion can quickly lead to loss of internal components andsubsequent dysfunction. In particular, the use of a positively-chargedemulsifying agent in the emulsion adds significantly to the activity ofthe emulsion, since it is attracted to the proteinaceous spore coat,where the positively charged oil-water interface denatures the proteinsand breaks down the spore's protective coating. If desired, sporicidalagents can also be included in the composition to increase theeffectiveness of the emulsion by taking advantage of theemulsion-damaged spore coat to penetrate into the spore and destroy themetabolically important components inside.

Still another aspect of the invention relates to added benefits to skin,including moisturizing and skin-softening properties associated withemulsions. Emollients, colors, and fragrances may be added to enhancethe cosmetic appeal of the product. Since emulsions include both waterand oil phases, a wide variety of such additives are possible withoutnegatively affecting the sporicidal activity. When applied to the skinas a leave-on product, emulsions can help extend protection for hoursbeyond the initial application. When an emulsion is formulated withincreased levels of non-volatile components, the sporicidal protectionof the composition may be extended for longer periods of time.

The key to spore resistance is the integrity of the spore coat and thelow water content of the spore protoplast. Upon activation andsubsequent germination, the spores become much more susceptible tosporicidal and bactericidal agents. Bactericidal agents such asalcohols, most quaternary ammonium compounds, phenols, hydrogenperoxide, iodine, triclocarban and triclosan exhibit very poorsporicidal activity per se. However, combining these agents with one ormore spore coat opening agents dramatically expands their efficacyagainst spores. Increased permeability of the spore coat allows theseagents to more readily penetrate the protective coating, allowing themaccess to vital sites within the spore. Therefore, the novel combinationof spore coat opening agents with these antimicrobial agents expandstheir traditional activities beyond vegetative cells to includesignificantly enhanced and expanded activity against spores.Applications for such combinations include enhanced antimicrobiallotions, handwashes, creams, gels, soaps, cleansers, first-aidantiseptics, rinse and no-rinse sanitizers, and the like applied to bothanimate and inanimate surfaces.

The present invention provides for safe and effective topical skin careformulations capable of killing microbiological spores, preventing theirsubsequent development into infectious, vegetative cells, and destroyingvegetative cells that result from outgrowth of spores. Furthermore, thenovel formulation is gentle to skin, providing a cosmetically pleasingfeel and assisting in gently moisturizing the skin, such that the userwill gain benefit from both the sporicidal activity and improved skinhealth through repetitive application. Certain embodiments of theinvention are also useful for cleaning and sterilizing hard surfaces.

Therefore, in one embodiment the present invention provides a germicidalcomposition with enhanced microbiocidal activities for killingmicrobiological spores, microbiological outgrowth from spores andvegetative microorganisms comprising at least one quaternary ammoniumcompound selected from the group consisting of a quaternary ammoniumcompound containing a C₁₈₋₃₀ alkyl substituent, benzethonium chloride,benzethonium bromide, benzethonium iodide, methylbenzethonium chloride,methylbenzethonium bromide, and methylbenzethonium iodide. Thequaternary ammonium compound is present in a concentration of about0.01% to 10%, or more preferably in a concentration of about 0.05% to8%, or in an optimal concentration of about 0.08% to 5%. The compositioncan be applied to healthy, infected, or wounded tissue such asepithelial tissues, skin, fingernails, toenails, hair, mammary glands,the perineal region, genitalia, rectum, and mucosal linings of the noseand/or the vaginal area in the form of a liquid spray, dipping solution,foam, lotion, cream, handwash, wet wipe, or the like, to prevent and/orprotect against the risk of infection. When applied to skin or aninanimate surface, the composition cleans the surface effectively,removing dirt, soil, oils, and the like.

In another embodiment, the present invention also provides a germicidalcomposition with enhanced microbiocidal activities for killingmicrobiological spores, microbiological outgrowth from spores, andvegetative microorganisms, containing at least one active microbiocidalagent and at least one spore coat opener. The combination of themicrobiocidal agent and spore coat opener is sporicidal. A preferredmicrobiocidal agent, which is also a sporicidal agent per se includes aquaternary ammonium halide compound selected from the group ofbenzethonium, methylbenzethonium, benzalkonium, a C₈₋₂₄ mono-alkylquaternary ammonium, a C₈₋₂₄ di-alkylquaternary ammonium chloride orbromide, or the like. Other quaternary ammonium quaternary ammoniumcompounds, which are microbiocidal but not effectively sporicidal alsocombine with spore coat openers to become sporicidal. Exemplary of suchquaternary ammonium compounds are simple quaternary ammonium compoundslike cetrimonium and benzalkonium halides.

The quaternary ammonium compound is present in a concentration of about0.05% to 10%, or more preferably in a concentration of about 0.08% to8%, or in an optimal concentration of about 0.08% to 5%.

Another example of a microbiocide that, in the presence of a spore coatopener, becomes a sporicidal agent is a phenolic compound chosen fromthe group consisting of phenol, para-chlorometaxylenol, cresol,resorcinol, hexylresorcinol, or the like. The phenolic compound ispresent in a concentration of about 0.01% to 10%, or more preferably ina concentration of about 0.01% to 5%, or in an optimal concentration ofabout 0.05% to 1%.

In addition, when combined with a spore coat opener, hydrogen peroxidecan be used as a sporicidal agent when present in a concentration ofabout 1% to 10%, or a more preferable concentration of about 1% to 5%,or an optimal concentration of about 2% to 4%.

Another microbiocide that, in the presence of a spore coat opener,becomes sporicidal includes iodine, in a concentration of about 0.1% to10%, or more preferably in a concentration of about 0.5% to 8%, or in anoptimal concentration of about 1% to 5%.

Furthermore, a compound selected from the group of triclocarban,triclosan, and the like, in a concentration of about 0.01% to 5%, ormore preferably in a concentration of about 0.05% to 3%, or in anoptimal concentration of about 0.05% to 2% becomes sporicidal in thepresence of a spore coat opener.

Suitable spore coat openers includes amino acids such as alanine,glycine, valine, leucine, isoleucine, proline, serine, threonine,methionine, cysteine, taurine, tyrosine, phenylalaine, tryptophan,asparagine, glutamine, aspartic acid, glutamic acid, lysine, arginine,histidine, or the like, including their free acid forms and their salts,wherein the amino acid or combinations of amino acids are present in aconcentration of about 0.0001% to 5%, or more preferably in aconcentration of about 0.001% to 2%, or in an optimal concentration ofabout 0.005% to 1%.

Another class of spore coat openers includes metal chelation agentschosen from the group consisting of EDTA, EGTA, picolinic acid,nicotinic acid, niacin, citric acid, ascorbic acid, glutamic acid,fulvic acid, and the like, including their free acid forms and theirsalts. The chelation agent is present in a concentration of about 0.001%to 5%, or more preferably in a concentration of about 0.01% to 2%, or inan optimal concentration of about 0.05% to 1%.

Further a reducing agent that is capable of reducing disulfide bondssuch as ascorbic acid, quinone, polyphenol, proanthocyanidin, or thelike, including their free acid forms and their salts can also be usedas spore coat openers. The reducing agent is present in a concentrationof about 0.001% to 2%, or more preferably in a concentration of about0.001% to 1%, or in an optimal concentration of about 0.005% to 0.5%.

The composition of the present invention optionally contains at leastone hydrophobic component chosen from the group consisting ofpetrolatum, mineral oil, dimethicone, synthetic or naturally occurringoils, and the like, wherein the hydrophobic component is present in aconcentration of about 0.1% to 70%, or more preferably in aconcentration of about 1% to 50%, or in an optimal concentration ofabout 1% to 35%.

The composition may also contain at least one emollient/solubilizer,chosen from the group consisting of selected from the group consistingof C₁₂₋₂₄ monohydric alcohols, C₂₋₁₈ monohydrohyric alcohol ethoxylates,C₈₋₂₄ alkylamine oxides, silicone copolyols, the quaternary ammoniumcompounds behentrimethylammonium, babassuamindopropalkonium,benzethonium, benzalkonium, a C₈₋₂₄ mono-alkylquaternary ammonium, alinear C₈₋₂₄ di-alkylquaternary ammonium and the like, combined withtheir respective anionic salt counterparts chosen from the group ofhalogens such as chloride, bromide, or iodide, methosulfate,ethosulfate, acetate, lactate, propionate, and the like. Theemollient/solubilizer is present in a concentration of about 0.1% to25%, or more preferably in a concentration of about 0.5% to 15%, or inan optimal concentration of about 1% to 10%.

Suitable emulsifiers for the present invention includepositively-charged, amphoteric, neutral or non-ionic surface-activeagents. The amphoteric or zwiterionic emulsifiers have both positive andnegative charges on the same emulsifier. A good example is a “betaine”,such as cocoamidopropyl betaine or behenyl betaine (amphotericsurfactants) or a “sultaine”, such as cocamidopropyl hydroxysultaine orerucamidopropyl hydroxysultaine. All of the positively-chargedantimicrobial quaternary ammonium compounds described in thisapplication also may serve a dual role in their respective formulationsas emulsifiers. In addition to these, other positively-chargedsurface-active agents may be utilized as emulsifiers, such ashydroxyethyl behenamidopropyl dimonium chloride. Example nonionicemulsifiers includes the following: sorbitan esters and ethoxylates suchas: sorbitan stearate, sorbitan oleate, sorbitan isostearate,polysorbate 20, polysorbate 60, polysorbate 80, PEG-20 sorbitanisostearate; ethoxylated modified glycerides such as: PEG-20 (almond orcorn) glycerides, PEG-60 (almond or corn) glycerides, PEG-45 palm kernelglycerides, PEG-6 caprylic/capric glycerides, PEG-7 glyceryl cocoate;Alkyl ethoxylates such as: Ceteth-20, PEG-30 caster oil, oleth-3,oleth-5, oleth-10, oleth-20, glycereth-31, steareth-2, steareth-10, andstereth-20; Carbohydrate esters such as: sucrose distearate, sucrosedistearate, sucrose cocoate, and the like; propoxylates such as: PPG-12butyl ether, PPG-10 butanediol, PPG-10 cetyl ether, PPG-3 myristylether; alkanolamides such as: cocamide DEA, lauramide DEA, stearamideDEA, and the like; amino oxides such as: behenamine oxide,cocamidopropylamine oxide, lauramine oxide, myristamine oxide,stearamine oxide; lanolin and lanolin derivatives and siliconecopolyols.

The present invention also provides a method for disinfecting surfacesand killing microbiological spores, microbiological outgrowth fromspores, vegetative microorganisms, viruses, molds, yeasts, and fungicomprising the step of applying a liquid to a surface, said liquidcontaining a composition of the present invention.

Since many modifications, variations and changes in detail can be madeto the described preferred embodiment of the invention, it is intendedthat all matters in the foregoing description and the following examplesare interpreted to illustrate, and not in any way to limit, theinvention.

EXAMPLE GROUP A

This type of sporicidal formulation contains one or more surface-activequaternary ammonium compounds, water, a polyhydric alcohol, one or morepreservatives, and if desired, additives to enhance the cosmetic aspectsof the formula such as fragrance, color, viscosity builders, emollients,and the like. Group A Examples: Sporicidal Liquid Formulations A1 A2 A3A4 A5 Distilled Water 98.16% 97.30% 97.68% 97.10% 96.58% Benzethoniumchloride, USP 0.20% 0.20% Stearalkonium chloride 0.20% Behentrimoniumchloride 0.05% Behentrimonium methosulfate 0.10% Benzalkonium chloride,80% 0.16% Cetrimonium chloride, 30% 0.50% 0.50% para-chlorometaxylenol0.20% 0.10% Glycerin 1.40% 1.00% 2.00% Propylene glycol 2.00% 2.00%Methylparaben 0.02% 0.02% 0.03% 0.02% 0.02% Propylparaben 0.02% 0.02%0.03% 0.02% 0.02% Cocoamidopropylbetaine, 30% 0.05% 0.10% 0.02%Ceteth-20 0.05% 0.05% 0.05% Squalane 0.01% 0.10% Aloe vera extract 0.01%0.01% 0.01% Vitamin E Acetate 0.10% 0.05% Fragrance & Preservatives q.s.q.s. q.s. q.s. q.s.

EXAMPLE GROUP B

These sporicidal formulations are similar to the Examples in Group A,but to which is added one or more water-insoluble hydrophobesappropriate for application to the skin with appropriate emollientsand/or solubilizers. Example Group B: Sporicidal Compositions withHydrophobes and Emollient/Solubilizers B1 B2 B3 B4 B5 Water Phase: Water68.0% 78.1% 82.7% 85.0% 73.7% Glycerin 2.5% 2.5% 2.5% Propylene glycol2.5% 2.5% 2.5% Benzethonium chloride 0.2% 0.2% 0.5% Cetrimethylammoniumchloride-30% 0.5% 0.1% para-chlorometaxylenol 0.1% 0.1% Aloe VeraExtract 0.1% 0.1% 0.1% Cucumber Extract 0.1% 0.1% Oil Phase: Mineral Oil11.4% 5.2% 4.4% 6.5% Petrolatum 6.6% 3.0% 2.6% Isopropyl Myristate 8.3%3.2% 3.9% Isoceryl Stearate 0.3% 4.3% 0.5% Dimethicone -350 0.8% 2.0%1.0% Stearalkonium chloride 4.0% 2.8% Ceteryl alcohol 6.5% 5.0% 3.9%4.1% 5.2% Behentrimonium chloride 2.1% Behentrimonium methosulfate 1.4%2.3% Babassuamidopropalkonium chloride 2.0% Vitamin E Acetate 0.1% 0.1%0.1% 0.1% 0.1% Preservatives and Fragrance: q.s. q.s. q.s. q.s. q.s.Mixing Directions:Mix ingredients for Water Phase and heat to 80° C. with mixing.Vlix ingredients for Oil Phase and heat to 80° C. with mixing in aseparate vessel.Combine Oil Phase and Water Phase with homogenization and/or agitation.Cool to below 50° C.; add preservatives and fragrance.

EXAMPLES GROUP C

These examples demonstrate novel compositions containing varioussporicidal agents in combination with spore coat opening agents. Group CExamples: Sporicidal Liquid Formulations with Spore Coat Opening AgentsC1 C2 C3 C4 C5 Distilled Water 98.06% 96.90% 97.34% 35.19% 96.00%Benzethonium chloride, USP 0.20% 0.20% 0.20% Stearalkonium chloride0.12% Behentrimonium chloride 0.10% Behentrimonium methosulfate 0.10%Benzalkonium chloride, 80% 0.20% 0.20% Cetyltrimethylammonium chloride,30% 0.50% 0.50% para-chlorometaxylenol 0.20% Ethyl alcohol, 95% 61.00%Glycerin 1.40% 1.40% 2.50% Propylene glycol 2.00% 2.00% Methylparaben0.02% 0.02% 0.02% 0.02% Propylparaben 0.02% 0.02% 0.02% 0.02%Cocoamidopropylbetaine, 30% 0.05% 0.05% Ceteth-20 0.05% 0.05% Isopropylmyristate 1.20% Squalane 0.01% 0.01% 0.01% Aloe vera extract 0.01% 0.01%Vitamin E Acetate 0.10% 0.10% Fragrance 0.09% 0.20% 0.09% 0.20% Kathon0.05% 0.05% 0.05% Sodium hydroxymethylglycinate 0.20% 0.20% Amino acidMixture 0.20% 0.01% 0.20% L-alanine 0.10% Calcium dipicolinate 0.10%0.05% Disodium EDTA 0.1% 0.01% Quinone 0.05% Proanthocyanidin 0.05%

EXAMPLES Groups D and E

Formulations such as taught in Examples A, B, and C are enhanced withrespect to sporicidal activity by combining one or more ingredients thatare known to trigger spore germination with hydrophobic components andappropriate emollients and solubilizers. These examples arerepresentative of the possible combinations of these components.Examples D&E: Sporicidal Emulsions Enhanced with Spore Coat OpeningAgents D1 E1 E2 E3 E4 Water Phase: Water 67.8% 77.8% 82.6% 84.9% 73.6%Glycerin 2.5% 2.5% 2.5% Propylene glycol 2.5% 2.5% 2.5% Benzethoniumchloride 0.2% 0.2% 0.5% Cetrimethylammonium chloride-30% 0.5% 0.1%para-chlorometaxylenol 0.1% 0.1% Aloe Vera Extract 0.1% 0.1% 0.1%Cucumber Extract 0.1% 0.1% Amino acid Mixture 0.3% L-alanine 0.1%Calcium dipicolinate 0.2% Disodium EDTA 0.1% Reducing Agent 0.1% OilPhase: Mineral Oil 11.4% 5.2% 4.4% 6.5% Petrolatum 6.6% 3.0% 2.6%Isopropyl Myristate 8.3% 3.2% 3.9% Isocetyl Stearate 0.3% 4.3% 0.5%Dimethicone -350 0.8% 2.0% 1.0% Stearalkonium chloride 4.0% 2.8% Ceterylalcohol 6.5% 5.0% 3.9% 4.1% 5.2% Behentrimonium chloride 2.1%Behentrimonium methosulfate 1.4% 2.3% Babassuamidopropalkonium chloride2.0% Vitamin E Acetate 0.1% 0.1% 0.1% 0.1% 0.1% Preservatives andFragrance: q.s. q.s. q.s. q.s. q.s.Mixing Directions:Mix ingredients for Water Phase and heat to 80° C. with mixing.Mix ingredients for Oil Phase and heat to 80° C. with mixing in separatevessels.Combine oil and water phases with homogenization and/or agitation.Cool to below 50° C., add preservatives and fragrance.Test Methods and Results:

Spore suspensions were prepared by streaking standard agar plates withBacillus subtilis and then incubating the plates for 48-72 hours at 37°C. The resulting bacteria and their spores were scraped from the platesand suspended by agitation in a 50% ethanol-water solution for fourhours in order to lyse the vegetative cells. The spores were collectedby centrifugation, washed three times and then suspended in 50%ethanol-water, resulting in a spore titer of 10⁹ spores/mL. Aliquots ofthis spore suspension were diluted 1:100 into representative embodimentsof the invention in order to demonstrate sporicidal activity. Aftertreatment, the spore suspensions were diluted in distilled water andduplicate aliquots from each dilution were pour-plated on tripticase soyagar. Total colony-forming units (cfu) were counted after incubation at37° C. for 24 hours. Sporicidal activity was directly proportional tothe reduction of cfu compared to the distilled water controls.

Test results from representative compositions are summarized in Table 1and the associated graph of the data (FIG. 2). In addition, test datafor exposure to 0.1% L-alanine in the absence of sporicidal agents isincluded. This demonstrates enhancement of the germination process byL-alanine as it helps to open the spore coat more effectively. TABLE 1Test results of Spore-inactivating Compositions vs. Exposure TimeTime(hrs) L-alanine A1 C1 B3 E2 0.00 100.0% 100.0% 100.0% 100.0% 100.0%0.25 106.0% 68.0% 35.0% 40.0% 21.0% 0.50 111.0% 49.0% 9.0% 28.0% 12.0%1.00 113.0% 19.0% 4.0% 8.0% 2.2% 4.00 116.0% 6.4% 5.2% 1.6% 0.1%

Similar experiments were also carried out against another type of spore.Clostridium sporogenes (ATCC# 7955) was chosen as a representative ofthe Clostridium genus for this test. Spores were exposed to 15-secondand 30-second exposures to formulation B3. When suspensions of thesespores were exposed to formula B3 (10⁷/mL final) for 15 seconds,reduction of 85% was observed. Exposure for 30 seconds under the sameconditions resulted in a reduction of 99.9%.

It is to be understood that the above-described arrangements are onlyillustrative of application of the principles of the present invention.Numerous modifications and alternative arrangements can be devisedwithout departing from the spirit and scope of the present invention.Thus, while the present invention has been shown in the drawings andfully described above with particularity and detail in connection withwhat is presently deemed to be the most practical and preferredembodiment(s) of the invention, it will be apparent to those of ordinaryskill in the art that numerous modifications, including, but not limitedto, variation in size, materials, shape, form, function and manner ofoperation, assembly and use may be made without departing from theprinciples and concepts set forth herein.

1. A germicidal composition with enhanced microbiocidal activities forkilling or disinfecting microbiological spores, microbiologicaloutgrowth from spores and vegetative microorganisms comprising 0.05% to10% of at least one quaternary ammonium compound selected from the groupconsisting of a quaternary ammonium compound containing a C₁₈₋₃₀ alkylsubstituent, benzethonium chloride, benzethonium bromide, benzethoniumiodide, methylbenzethonium chloride, methylbenzethonium bromide andmethylbenzethonium iodide.
 2. A germicidal composition with enhancedmicrobiocidal activities for killing or inhibiting microbiologicalspores, microbiological outgrowth from spores and vegetativemicroorganisms comprising: at least one active sporicidal agent selectedfrom the group consisting of: (1) a quaternary ammonium compoundselected from the group consisting of benzethonium chloride or bromide,methylbenzethonium chloride or bromide, benzalkonium chloride orbromide, a C₈₋₂₄ mono-alkyl quaternary ammonium chloride or bromide, anda C₈₋₂₄ di-alkylquaternary ammonium chloride or bromide; (2) a phenoliccompound selected from the group consisting of phenol,para-chlorometaxylenol, cresol and hexylresorcinol; (3) a compoundselected from the group consisting of triclocarban, triclosan; amonohydric alcohol, hydrogen peroxide and iodine; and at least one sporecoat opener selected from the group consisting of: (1) an amino acid ora salt form of an amino acid; (2) a metal chelation agent selected fromthe group consisting of EDTA, EGTA, picolinic acid, niacin, citric acid,ascorbic acid, glutamic acid, and fulvic acid; and (3) a reducing agentselected from the group consisting of ascorbic acid, quinone,polyphenol, proanthocyanidin, and their salts.
 3. The germicidalcomposition of claim 2 wherein the sporicidal agent is at least onequaternary ammonium compound selected from the group consisting ofbenzethonium chloride or bromide, methylbenzethonium chloride orbromide, benzalkonium chloride or bromide, a C₈₋₂₄ mono-alkyl quaternaryammonium chloride or bromide, and a C₈₋₂₄ di-alkylquaternary ammoniumchloride or bromide, and the spore coat opener is a member selected fromthe group consisting of an amino acid, picolinic acid, niacin, citricacid, ascorbic acid, quinone, polyphenol, proanthocyanidin and theirsalts.
 4. The germicidal composition of claim 3 wherein the sporicidalagent is at least one member selected from the group consisting ofphenol, para-chlorometaxylenol, cresol, hexylresorcinol, triclocarban, amonohydric alcohol, hydrogen peroxide and iodine.
 5. A germicidalcomposition with enhanced microbiocidal activities for killing ordisinfecting microbiological spores, microbiological outgrowth fromspores and vegetative microorganisms comprising: at least one activesporicidal agent selected from the group consisting of: (1) a quaternaryammonium compound selected from the group consisting of benzethoniumchloride or bromide, methylbenzethonium chloride or bromide,benzalkonium chloride or bromide, a C₈₋₂₄ mono-alkyl quaternary ammoniumchloride or bromide, and a C₈₋₂₄ di-alkylquaternary ammonium chloride orbromide; (2) a phenolic compound selected from the group consisting ofphenol, para-chlorometaxylenol, cresol and hexylresorcinol; and (3) acompound selected from the group consisting of triclocarban, triclosan,a monohydric alcohol, hydrogen peroxide, and iodine; at least one sporecoat opener selected from the group consisting of: (1) an amino acid ora salt form of an amino acid; (2) a metal chelation agent selected fromthe group consisting of EDTA, EGTA, picolinic acid, niacin, citric acid,ascorbic acid, glutamic acid, and fulvic acid, and their salts; and (3)a reducing agent selected from the group consisting of ascorbic acid,quinone, polyphenol, proanthocyanidin, and their salts; at least onehydrophobic component selected from the group consisting of petrolatum,mineral oil, dimethicone, and synthetic or naturally occurring oils; andat least one emollient or solubilizer selected from the group consistingof C₁₂₋₂₄ monohydyric alcohols, C₂₋₁₈ monohydrohyric alcoholethoxylates, C₈₋₂₄ alkylamine oxides, silicone copolyols,behentrimethylammonium methosulfate, behentrimethylammonium ethosulfate,behentrimethylammonium chloride, behentrimethylammonium bromide,babassuamindopropalkonium chloride, babassuamindopropalkonium bromide,benzethonium chloride or bromide, benzalkonium chloride or bromide, aC₈₋₂₄ mono-alkylquaternary ammonium chloride or bromide, and a C₈₋₂₄di-alkylquaternary ammonium chloride or bromide.
 6. The germicidalcomposition of claim 5 wherein the sporicidal agent is at least onequaternary ammonium compound selected from the group consisting ofbenzethonium chloride or bromide, methylbenzethonium chloride orbromide, benzalkonium chloride or bromide, a C₈₋₂₄ mono-alkyl quaternaryammonium chloride or bromide, and a C₈₋₂₄ di-alkylquaternary ammoniumchloride or bromide, and the spore coat opener is a member selected fromthe group consisting of an amino acid, picolinic acid, niacin, citricacid, ascorbic acid, glutamic acid, fulvic acid, quinone, polyphenol,proanthocyanidin and their salts.
 7. The germicidal composition of claim5 wherein the sporicidal agent is at least one member selected from thegroup consisting of phenol, para-chlorometaxylenol, cresol,hexylresorcinol, triclocarban, triclosan; a monohydric alcohol, hydrogenperoxide and iodine.
 8. A method for disinfecting surfaces and killingmicrobiological spores, microbiological outgrowth from spores, andvegetative microorganisms comprising the step of applying thecomposition of claim 1 to a surface.
 9. The method of claim 8, whereinthe surface is a member selected from the group consisting of epithelialtissues, skin, fingernails, toenails, hair, mammary glands, perinealregion, genitalia, rectum and mucosal linings.
 10. The method of claim8, wherein the surface is an inanimate surface.
 11. The method of claim8, wherein the composition is applied to the surface in the form of aspray, dipping solution, foam, lotion, cream, handwash, or wet wipe. 12.A method for disinfecting surfaces and killing microbiological spores,microbiological outgrowth from spores, and vegetative microorganismscomprising the step of applying the composition of claim 5 to a surface.13. The method of claim 12, wherein the surface is a member selectedfrom the group consisting of epithelial tissues, skin, fingernails,toenails, hair, mammary glands, perineal region, genitalia, rectum andmucosal linings.
 14. The method of claim 12, wherein the surface is aninanimate surface.
 15. The method of claim 12, wherein the compositionis applied to the surface in the form of a spray, dipping solution,foam, lotion, cream, handwash, or wet wipe.